The goal of this proposal is to understand how a novel DNA polymerase, Pol K, functions to couple replication fork passage to establishment of cohesion between newly formed sister chromatids. We have recently determined that po1 K (encoded by TRF4 and TRF5) is essential for both replication and cohesion (Wang et al. (2000) Science 289, 774-779). These observations open the door to further mechanistic insight into how cohesion establishment is accomplished, how it is coupled to replication fork passage and perhaps to the nature of the bond between sister chromatids. The mechanism by which cohesion is formed during replication is crucial for the maintenance of genomic integrity, but nonetheless remains a fundamental unanswered question in cell biology. We hypothesize that POI K is an essential component of the replication fork in all eukaryotes whose function is to replicate cohesion sites specifically and, thereby, set up the cohesed state between chromatids. We further hypothesize that an interaction between po1 K and the non-polymerase C-terminal domain of pol E about occurs and that this interactions is necessary forpol K to act as an S-phase checkpoint sensor. Experiments designed to test our hypothesis are outlined below.